Staphylococcaceae

STAPHYLOCOCCACEAE

Instructor

Dr. Maytham Ihsan

Ph.D Vet Microbiology

STAPHYLOCOCCACEAE

Genus: Staphylococcus Staphylococci genera, are Gram‐positive cocci, approximately 1 μm in diameter, that tend to occur in irregular clusters resembling bunches of grapes. Staphylococcus species occur as commensals on skin and mucous membranes; some may act as opportunistic pathogens causing pyogenic infections. Most staphylococci are facultative anaerobes and catalase‐ positive. They are non‐motile and oxidase‐negative and do not form spores. Two species, S. aureus subsp. anaerobius and S. saccharolyticus, are anaerobic and catalase‐negative. Micrococcus spp. (packs of tetrads), are positive to modified oxidase test. A total of 45 species of Staphylococcus have been described to date, seven of which are coagulase‐positive or coagulase‐ variable species. The coagulase‐positive S. aureus subsp. aureus (referred to as S. aureus), S. pseudintermedius, and the coagulase‐ variable S. hyicus are important pathogens of domestic animals. Staphylococcus intermedius it is now belong to the species S. pseudintermedius. Coagulase production correlates with pathogenicity. Although coagulase‐ negative staphylococci are usually of low virulence, some occasionally cause disease in animals and humans like S. epidermidis .

Staphylococci ‐ Natural Habitat • Non‐pathogenic staphylococci ‐ normal flora of skin

• Pathogenic staphylococci ‐ skin and mucous membranes (salt tolerant) • Survive in environment • Opportunistic infections: Wounds / teat damage (mastitis) / invade tick bites / (tick pyaemia) / pyoderma / more serious life‐threatening bacteraemia & septicaemia.

Differentiation of Gram Positive Cocci

Growth & Culture Characteristics • Staphylococci grow readily on most bacteriologic media under aerobic or microaerophilic condions at 37°C but form pigment best at room temperature (20–25°C). • Colonies on solid media are round, smooth, raised, and glistening. • S. aureus usually forms deep golden yellow colonies while S epidermidis colonies usually are gray to white on primary isolation; many colonies develop pigment only upon prolonged incubation. • S. aureus isolated from human & cow produced golden‐yellow discoloration, while those from dog were white. 3

• No pigment is produced anaerobically or in broth. • Are relavely resistant to drying, heat (they withstand 50°C for 30 minutes), and 9% sodium chloride but are readily inhibited by certain chemicals (eg, 3% hexachlorophene). • Tolerate bile up to 40%. • Produce a wide range of Exoenzymes & Exotoxins, playing an important role in their virulence & pathogensity. • Staphylococci are variably susceptible to many antimicrobial drugs. • Resistance to several antimicrobial drugs like β‐Lactamase production made them resistant to Penicillins. • Methicillin‐Resistant S aureus (MRSA) , Resistance to methicillin, nafcillin and to oxacillin. Most antimicrobial agents resistance properties carried via plasmid mainly by conjugation. Methicillin‐resistant staphylococcal infections in animals Infection with MRSA has been a major problem in human hospitals for many years and have become a major problem in veterinary medicine and animal production in the last decade. Transmission of infection between pets and humans, including veterinary personnel, and between horses and humans has been reported. Dogs & horses are widely infected with MRSA like: wound infections, surgical site infections, pyoderma, otitis and urinary tract infections.

Virulence factors: Including Exoenzymes & Exotoxins, of Staphylococcus aureus and their pathogenic effects Virulence factor Pathogenic effects Coagulase Conversion of fibrinogen to fibrin. Fibrin deposition may shield staphylococci from phagocytic cells Clumping Factor A cell wall bound, is responsible for adherence of the organisms to fibrinogen and fibrin. When mixed with plasma, S aureus forms clumps. Lipase, esterases, Enzymes which contribute to tissue destruction and virulence elastase, staphylokinase, deoxyribonuclease, hyaluronidase, phospholipase Catalase Converts hydrogen peroxide into water and oxygen. The staphylococci, are positive, whereas the streptococci, are negative. Protein A Surface component which binds Fc portion of IgG and inhibits opsonization Leukocidin Cytolytic destruction of phagocytes of some animal species Panton‐Valentine L Composed of two components S & F (PVL) Alpha‐toxin (alpha‐ The major toxin in gangrenous mastitis. It causes spasm of haemolysin) smooth muscle and is necrotizing and potentially lethal Beta‐toxin (beta‐ A sphingomyelinase which damages cell membranes haemolysin) Exfoliative toxins Proteases which contribute to skin lesion development in humans, dogs and pigs Staphylokinase Cause fibrinolysis Enterotoxins Heat‐stable toxins associated with staphylococcal food poisoning in humans T oxic shock Induce excessive lymphokine production, resulting in tissue syndrome toxins (T damage. Bovine and human strains of S. aureus produce T SST ‐1. SST ) Sheep and goat strains produce a variant of this toxin. T SST ‐1 has superantigen activity

Antigenic Structure ‐ Peptidoglycan , a polysaccharide polymer containing linked subunits, provides the rigid exoskeleton of the cell wall. Peptidoglycan is destroyed by strong acid or exposure to lysozyme. It is important in the pathogenesis of infection: 1‐ It induces production of interleukin‐1 (endogenous pyrogen) ; 2‐ Opsonic antibodies by monocytes; 3‐ A chemoattractant for polymorphonuclear leukocytes; 4‐ Have endotoxin‐like activity and 5‐ Activate complement. ‐ Teichoic acids , which are polymers of polyribitol–phosphate, are cross‐linked to the peptidoglycan and can be antigenic. ‐ Protein A , is a cell wall component of S. aureus strains and is a bacterial surface protein that has been characterized among a group of adhesins.Protein A binds to the Fc portion of IgG molecules except IgG3.

Diseases Caused by Staphylococci Infection

Diagnosis • Specimens: Pus, blood, urine & exudates divided into two parts, one to be stained by Gram’s stain & the other part cultured on nutrient agar, ox or sheep blood agar along with MacConkey agar.

• Selecve media used for culvaon like Staph 110, Mannitol Salt Agar (MSA), Baird‐Parker Agar with Egg Yolk or with Tellurite from food. Azide blood agar or Clositin‐Naldixic Acid Blood Agar to isolate Stapylocooi from feces & sewage. • Colonial morphology & pigmentation. Milk agar used for enhancement of pigment production with addition of Glycerol. • Haemolysis on blood agar (α, β,γ & δ). S. aureus and S. pseudintermedius usually produce both alpha‐haemolysin and beta‐haemolysin with a characterisc double zone of haemolysis; 1st narrow zone Alfa & 2nd wide of Beta. Isolates of S. hyicus are non‐haemolytic. • Coagulase production clot pasma, done by: 1‐ Slide Coagulase: For detection of both bound coagulase & clumping factor, but mostly gave false positive. 2‐ Tube Coagulase: For detection of bound coagulase & used as confirmatory to slide coagulase. Rabbit plasma mixed with isolates used with both tests. • Biochemical Profile: Catalase, Coagulase, Dnase, Protein A, Phosphatase, CHO fermentation and others. Many manufacturers developed ready to use kits called biochemical identification system like: API STAP & RapID STAPH. Purple Agar: Contains 1% maltose. The indicator is Bromocresol purple for differentiation of S. aureus from S. pseudintermedius & S. intermedius • Antimicrobial susceptibility tests. • Molecular typing: PCR‐based techniques used widely recently for detection & genotyping of Staphylococci with gene sequencing for genetic barcoding of a defined target gene.

• Phage Typing: Used for many years in epidemiological investigations such as those relating to outbreaks of staphylococcal food poisoning in humans and animals. Specific type for each species result in plaques formation.